Metabolic Engineering of Cancer for Selective Immunotargeting Cancer is one of the most common and fatal diseases and, in general, it is difficult to cure. Among the different strategies probed for cancer therapy, treating cancer patients with cancer vaccines or cancer-targeting antibodies is especially attractive as the human immune system can be extremely effective and selective to eliminate tumors in the human body. In the development of cancer vaccines used for patient immunization or antibody preparation, the abnormal glycans expressed by cancer cells, called tumor-associated carbohydrate antigens (TACAs), are valuable molecular targets, as they are abundant, exposed, and conserved on the cancer cell surface. However, the problem is that TACAs are usually poorly immunogenic or tolerated by the patients' immune system, which has severely hindered the delivery of functional TACA-based cancer vaccines or immunotherapies. To address the problem and develop effective cancer cures, a new immunotherapeutic strategy was proposed. First, animals (or patients) are immunized with a vaccine made of an unnatural TACA derivative to elicit a specific immune response. Next, the animals are treated with a similarly modified monosaccharide to bioengineer tumor cell expression of the unnatural TACA derivative (cancer cell glycoengineering). Then, the trained immune system will specifically recognize and kill the glycoengineered tumors. The new immunotherapy can also be realized with a TACA derivative-specific antibody, instead of a vaccine, for the treatment. For the strategy to work, it has to mee two conditions, namely, a potent vaccine that can be used for patient immunization or antibody preparation and an effective method for cancer cell glycoengineering. It has been proved that the latter can exploit the flexible biosynthetic pathways for glycans. To create potent and reliabl TACA-based vaccines, a new vaccine strategy is proposed here, namely, to have TACAs linked to a bacterial monophosphoryl lipid A (MPLA). The hypothesis is that MPLA can act as a powerful vaccine carrier and built-in adjuvant to formulate fully synthetic, self-adjuvanting, structurally defined, readily reproducible, and robust glycoconjugate vaccines. This proposal aims to: (1) study the structure-activity relationship of MPLAs and identify new, potent vaccine carriers and adjuvants, (2) prepare and study the immunological properties of TACA-MPLA conjugates and identify the proper vaccines for cancer immunotherapy, and (3) use the new TACA-MPLA conjugates as vaccines for active and passive immunotherapy of cancer such as melanoma and breast or colon cancer by the above therapeutic strategy. One innovation of this project is the use of MPLA as a carrier and adjuvant for fully synthetic self-adjuvanting carbohydrate vaccine development and related studies. Another innovation is the combination MPLA conjugate vaccine with cell glycoengineering for cancer therapy. This combination will solve the immunotolerance problem of TACAs, a central issue in cancer immunology, and help develop functional cures for various tumors. The new vaccine strategy will be applicable to other vaccine design as well. Thus, both strategies should be widely useful, and this research program should be of general significance and have a broad impact on cancer research.